The use of integrating spheres or multi-pass cells for quantifying the optical density of a fluid is prolific; however, in most of these applications, the fluid being examined is disposed in a sample container (e.g. a cuvette) and placed inside or against a porthole manufactured in the side of the sphere of cell. A radiation source, typically a halogen, mercury, or deuterium lamp, is used to irradiate the fluid inside the sample container. Some light is absorbed by the fluid and the remainder is scattered into the interior of the sphere or cell. After being multiply scattered, the remaining light exits the sphere or cell through a porthole and is detected by a spectrometer or the like. The path length in this type of measurement is determined by the dimensions of the cuvette, and is not enhanced by multiple scattering inside the sphere or cell. This is a common methodology in the field of absorption spectroscopy, where the amount of light absorbed can be correlated to the concentration of a molecule in the fluid. In this methodology, attenuation due to light absorption can be separated from attenuation due to light scattering through the integrating sphere or multi-pass cell; however, there is no enhancement of absorption coefficient measurement, as the path length is determined strictly by the cuvette dimensions. Thus, what are still needed in the art are improved systems and methods for determining the absorption coefficient and the optical density of a fluid as they relate to the wavelength of incident radiation.
Further, nucleic acid quantitation is used prolifically to determine the presence and the concentration of deoxyribonucleic acid (DNA) and ribonucleic acid (RNA) in a sample. There are several methodologies used to measure such concentrations. When relatively low concentrations of DNA are present, fluorescent dyes are used that bind to the nucleic acid and the resulting fluorescent intensity is compared to control samples. This method can be more time consuming than other methods, but is more accurate at relatively low sample concentrations. The Slot-Blot technique can also be used for relatively low sample concentrations, but requires adding a hybridizing agent and relies on luminescence measurement. Absorption spectroscopy is a commonly used method whereby the sample absorbance at 260 nm is correlated to the concentration and the sample absorbance at 280 nm. It may also be used to ascertain sample purity or contamination of a protein sample by DNA as compared to the 260 nm absorption. This method is defined by the Beer-Lambert Law, where absorption is a function of the path length of the sample. Because, as described above, the samples of interest are typically in held small cuvettes (i.e. about 1-cm path length) or microplates, they must have sufficient DNA concentrations to allow for measurement, or, be mechanically concentrated prior to sampling (as required for a nanodrop spectrophotometer). If the path length of the sample can be increased, then the level of sensitivity of the measurement can be increased; alternatively, smaller sample volumes can be used for measurement. Thus, what are still needed in the art are improved systems and methods for determining the presence and the concentration of DNA and RNA in a sample by exploiting increased path length.